Activity of a Carboxyl-Terminal Truncated Form of Catechol 2,3-Dioxygenase fromPlanococcussp. S5

Abstract

Catechol 2,3-dioxygenases (C23Os, E.C.1.13.12.2) are two domain enzymes that catalyze degradation of monoaromatic hydrocarbons. The catalytically active C-domain of all known C23Os comprises ferrous ion ligands as well as residues forming active site pocket. The aim of this work was to examine and discuss the effect of nonsense mutation at position 289 on the activity of catechol 2,3-dioxygenase from Planococcus strain. Although the mutant C23O showed the same optimal temperature for activity as the wild-type protein (35°C), it exhibited activity slightly more tolerant to alkaline pH. Mutant enzyme exhibited also higher affinity to catechol as a substrate. Its K m (66.17 µM) was approximately 30% lower than that of wild-type enzyme. Interestingly, removal of the C-terminal residues resulted in 1.5- to 1.8-fold (P < 0.05) increase in the activity of C23OB61 against 4-methylcatechol and 4-chlorocatechol, respectively, while towards catechol the activity of the protein dropped to about 80% of that of the wild-type enzyme. The results obtained may facilitate the engineering of the C23O for application in the bioremediation of polluted areas.

A novel Bacillus ligniniphilus catechol 2,3-dioxygenase shows unique substrate preference and metal requirement

“…SI3 A) consistent with extradiol cleavage of 3-ethylcatechol by C23O enzymes as reported by literature. When cleavage products were detectable, the specific λmax for other tested compounds all agreed with literature reported wavelengths 45 – 47 confirming BLC23O enzymatic activities. The other tested substrates did not show visible peak or noticeable change in spectra [Fig.…”

“…The reactions were initiated by adding various substrate concentrations of 0.2–1.2 mM 3-methylcatechol or 0.05–1.0 mM 3-ethylcatechol. The activity of BLC23O was measured by the increase of cleavage product formation at OD 388 nm (3-methylcatechol) or 390 nm (3-ethylcatechol) every 30 s for 30 min in a 96 well UV-Star microplate (Greiner Bio-one) using a spectrophotometer (SpectraMax M5, Molecular Devices, San Jose, CA, USA) 45 . The initial linear slopes were used to determine the kinetic parameters of BLC23O.…”

Catechol 2,3‐dioxygenase from a new phenolic compound degrader Thauera sp. K11: purification and biochemical characterization

“…According to the coordination metal ion at the catalytic center, C23Os are classified into three groups: Fe‐C23O, Mn‐C23O, and Mg‐C23O . To determine the type of C23O from Thauera sp.…”